U.S. patent number 7,205,531 [Application Number 10/991,123] was granted by the patent office on 2007-04-17 for sample information measuring method and scanning confocal microscope.
This patent grant is currently assigned to Olympus Corporation. Invention is credited to Wataru Nagata, Hideo Watanabe.
United States Patent |
7,205,531 |
Watanabe , et al. |
April 17, 2007 |
Sample information measuring method and scanning confocal
microscope
Abstract
When irradiating a sample with light from a light source through
an object lens, discretely changing a relative position between a
beam condensing position of the object lens and the sample in an
optical axis direction of the converging beam, obtaining light
intensity information from the sample at each relative position,
extracting plural pieces of light intensity information from a
light intensity information group, estimating a maximum value on a
change curve adaptive to the plural pieces of extracted light
intensity information and the relative position for the maximum
value, and obtaining the estimated maximum value of the light
intensity information and relative position as brightness
information and height information, these information about the
sample can be continuously obtained by discretely performing an
iterative operation on the relative position between a beam
condensing position of the object lens and the sample in an optical
axis direction of the converging beam.
Inventors: |
Watanabe; Hideo (Tokyo,
JP), Nagata; Wataru (Tokyo, JP) |
Assignee: |
Olympus Corporation (Tokyo,
JP)
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Family
ID: |
34742005 |
Appl.
No.: |
10/991,123 |
Filed: |
November 17, 2004 |
Prior Publication Data
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Document
Identifier |
Publication Date |
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US 20050161592 A1 |
Jul 28, 2005 |
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Foreign Application Priority Data
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Nov 21, 2003 [JP] |
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2003-391642 |
Nov 12, 2004 [JP] |
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2004-328368 |
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Current U.S.
Class: |
250/234;
250/201.3; 250/548; 356/609; 359/383 |
Current CPC
Class: |
G02B
21/006 (20130101) |
Current International
Class: |
H01J
3/14 (20060101); G02B 21/00 (20060101) |
Field of
Search: |
;250/201.3,559.22,234,221,548 ;359/368,383 ;356/609 |
References Cited
[Referenced By]
U.S. Patent Documents
Foreign Patent Documents
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9-68413 |
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Mar 1997 |
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JP |
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9-113235 |
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May 1997 |
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JP |
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11-264933 |
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Sep 1999 |
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JP |
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Primary Examiner: Pyo; Kevin
Attorney, Agent or Firm: Scully, Scott, Murphy & Presser
PC
Claims
What is claimed is:
1. A sample information measuring method, comprising: irradiating a
sample with light from a light source through an object lens;
discretely changing a relative position between a beam condensing
position of the object lens and the sample in an optical axis
direction of the converging beam; obtaining light intensity
information from the sample at each relative position; extracting
plural pieces of light intensity information from a light intensity
information group; estimating a maximum value on a change curve
adaptive to the plural pieces of extracted light intensity
information and the relative position for the maximum value; and
obtaining the estimated maximum value of the light intensity
information and relative position as brightness information and
height information, wherein the brightness information and height
information about the sample can be continuously obtained by
discretely performing an iterative operation on the relative
position between a beam condensing position of the object lens and
the sample in an optical axis direction of the converging beam, and
processing light intensity information from the sample at each
relative position obtained in a time between an inverse operation
of the moving mechanism and a next inverse operation during the
iterative operation.
2. The method according to claim 1, wherein three-dimensional image
is obtained with high-precision with equal or smaller travel
interval (step) of discretely changing the converging beam in an
optical axis direction when an instruction is issued to fetch an
image for a measurement mode in which the sample is measured with
high precision when the brightness information and height
information about the sample are continuously obtained by
discretely performing an iterative operation on the relative
position between a beam condensing position of the object lens and
the sample in an optical axis direction of the converging beam, as
compared with a time when the brightness information and height
information about the sample is continuously obtained by the
iterative operation.
3. The method according to claim 1, wherein three-dimensional image
is obtained with high precision with equal or smaller
two-dimensional scanning interval (step) of scanning a beam
condensing position of an object lens relative to the sample in a
direction perpendicular to an optical axis of the converging beam
when an instruction is issued to fetch an image for a measurement
mode in which the sample is measured with high precision when the
brightness information and height information about the sample are
continuously obtained by discretely performing an iterative
operation on the relative position between a beam condensing
position of the object lens and the sample in an optical axis
direction of the converging beam, as compared with a time when the
brightness information and height information about the sample is
continuously obtained by the iterative operation.
4. The method according to claim 1, wherein a shape of the sample
is continuously updated and displayed according to the brightness
information and height information about the continuously obtained
sample by the iterative operation.
5. The method according to claim 4, wherein a measurement position
is specified on a sample shape continuously updated and displayed
by the iterative operation, and the sample shape is measured based
on the specified measurement position.
6. A scanning confocal microscope, comprising: an object lens
converging light from a light source in a sample; a moving
mechanism relatively moving a beam condensing position of the
object lens with a position of the sample in an optical axis
direction of the converging beam; a confocal diaphragm arranged in
a position conjugate with a beam condensing position of the object
lens; and an photodetector detecting intensity of light passing
through the confocal diaphragm; a unit discretely performing an
iterative operation on a relative position between a beam
condensing position of the object lens to the sample in an optical
axis direction of the converging beam by the moving mechanism; a
unit obtaining light intensity information from the sample at each
relative position; a unit extracting plural pieces of light
intensity information from a light intensity information group; a
unit estimating a maximum value on a change curve adaptive to the
plural pieces of extracted light intensity information, and the
relative position for the maximum value; and a brightness and
height information arithmetic unit obtaining the estimated maximum
value of light intensity information and relative position as
brightness information and height information, wherein the
brightness and height information arithmetic unit processes light
intensity information from the sample at each relative position
obtained in a time between an inverse operation of the moving
mechanism and a next inverse operation, and continuously obtains
the brightness information and height information in
synchronization with an iterative operation of the moving
mechanism.
7. The microscope according to claim 6, further comprising a
three-dimensional image data acquisition instruction unit fetching
an image for a measurement mode in which the sample is measured
with high precision, wherein the brightness and height information
arithmetic unit obtains three-dimensional image data with a high
precision at equal or shorter travel intervals (steps) of
discretely changing the converging beam in an optical axis
direction when the three-dimensional image data acquisition
instruction unit is specified when the brightness information and
height information about the sample are continuously obtained by
discretely performing an iterative operation on the relative
position between a beam condensing position of the object lens and
the sample in an optical axis direction of the converging beam, as
compared with a time when the brightness information and height
information about the sample are continuously obtained by the
iterative operation.
8. The microscope according to claim 6, further comprising: a
two-dimensional scanning unit scanning in a direction perpendicular
to an optical axis of the converging beam; and a three-dimensional
image data acquisition instruction unit fetching an image for a
measurement mode in which the sample is measured with high
precision, wherein the brightness and height information arithmetic
unit obtains three-dimensional image data with high precision at
equal or shorter two-dimensional scanning intervals (steps) of
performing scanning in a direction perpendicular to an optical axis
of the converging beam when the three-dimensional image data
acquisition instruction unit is specified when the brightness
information and height information about the sample are
continuously obtained by discretely performing an iterative
operation on the relative position between a beam condensing
position of the object lens and the sample in an optical axis
direction of the converging beam, as compared with a time when the
brightness information and height information about the sample are
continuously obtained by the iterative operation.
9. The microscope according to claim 6, further comprising a
display unit continuously updating and simultaneously displaying a
shape of the sample according to the brightness information and
height information about the continuously obtained sample.
10. The microscope according to claim 9, further comprising: a
measurement position designation unit designating a measurement
position on a sample shape continuously updated and displayed; and
a sample shape measurement unit measuring the sample shape based on
the measurement position designated by the measurement position
designation unit.
11. A sample information measuring method, comprising: irradiating
a sample with light from a light source through an object lens;
discretely changing a relative position between a beam condensing
position of the object lens and the sample in an optical axis
direction of the converging beam; obtaining light intensity
information from the sample at each relative position; extracting
plural pieces of light intensity information from a light intensity
information group; estimating a maximum value on a change curve
adaptive to the plural pieces of extracted light intensity
information and the relative position for the maximum value; and
obtaining the estimated maximum value of the light intensity
information and relative position as brightness information and
height information, wherein: the brightness information and height
information about the sample can be continuously obtained by
discretely performing an iterative operation on the relative
position between a beam condensing position of the object lens and
the sample in an optical axis direction of the converging beam and
processing light intensity information from the sample at each
relative position obtained in a time between an inverse operation
of the moving mechanism and a next inverse operation during the
iterative operation; and a 3D image of the sample shape generated
according to the obtained height information and the brightness
information are continuously updated and displayed on a same
screen.
12. The method according to claim 11, wherein a confocal image is
generated according to the brightness information, and is
continuously updated and displayed with the 3D image on a same
screen by the iterative operation.
13. The method according to claim 12, wherein the confocal image is
an extend image.
14. The method according to claim 11, wherein the 3D image is
updated and displayed each time the shape is obtained; and the
brightness information displayed with the 3D image is updated and
displayed each time the brightness information is obtained.
15. The method according to claim 11, wherein a timing of updating
an image is reported on a display screen for display of an image
while continuously updating the image by the iterative
operation.
16. A scanning confocal microscope, comprising: an object lens
converging light from a light source in a sample; a moving
mechanism relatively moving a beam condensing position of the
object lens with a position of the sample in an optical axis
direction of the converging beam; a confocal diaphragm arranged in
a position conjugate with a beam condensing position of the object
lens; and an photodetector detecting intensity of light passing
through the confocal diaphragm; a unit discretely performing an
iterative operation on a relative position between a beam
condensing position of the object lens to the sample in an optical
axis direction of the converging beam by the moving mechanism; a
unit obtaining light intensity information from the sample at each
relative position; a unit extracting plural pieces of light
intensity information from a light intensity information group; a
unit estimating a maximum value on a change curve adaptive to the
plural pieces of extracted light intensity information, and the
relative position for the maximum value; a brightness and height
information arithmetic unit obtaining the estimated maximum value
of light intensity information and relative position as brightness
information and height information, processing light intensity
information from the sample at each relative position obtained in a
time between an inverse operation of the moving mechanism and a
next inverse operation, and continuously obtaining the brightness
information and height information in synchronization with an
iterative operation of the moving mechanism; and a display unit
generating a 3D image of the sample shape according to the
brightness information and height information about the
continuously obtained sample, and continuously updating and
simultaneously displaying the image with the brightness information
on a same screen.
17. The microscope according to claim 16, wherein the display unit
generates a confocal image according to the brightness information,
and continuously updating and simultaneously displaying the image
with the 3D image on a same screen by the iterative operation.
18. The microscope according to claim 17, wherein the confocal
image is an extend image.
19. The microscope according to claim 16, wherein the display unit
updates and simultaneously displays the 3D image each time the
shape is obtained; and the brightness information is updated and
simultaneously displayed each time the brightness information is
obtained.
20. The microscope according to claim 16, wherein the display unit
comprises an image update and display unit displaying an image at
an update timing of the image.
21. The method according to claim 5, wherein a profile of the
sample at a measurement position specified in a shape of the sample
is displayed, and the shape of the sample is measured on the
profile.
22. The microscope according to claim 10, wherein a profile of the
sample at a measurement position in a shape of the sample
designated by the measurement position designation unit is
displayed, and the shape of the sample is measured on the
profile.
23. A sample information measuring method, the method comprising:
irradiating a sample with light from a light source through an
object lens; discretely moving a relative position between a beam
condensing position of the object lens and the sample in an optical
axis direction of the converging beam within a desired Z scanning
area; obtaining light intensity information from the sample at each
relative position; extracting plural pieces of light intensity
information from a light intensity information group; estimating a
maximum value on a change curve adaptive to the plural pieces of
extracted light intensity information and the relative position for
the maximum value; and obtaining the estimated maximum value of the
light intensity information and relative position as brightness
information and height information; wherein the brightness
information and height information about the sample can be
continuously obtained by discretely performing an iterative
operation on the relative position between a beam condensing
position of the object lens and the sample from a lower limit
position to an upper limit position and from an upper limit
position to the lower limit position of the Z scanning area in an
optical axis direction of the converging beam, and a shape of the
sample is continuously updated and displayed according to the
brightness information and height information about the
continuously obtained sample by the iterative operation.
24. The method according to claim 23, wherein a three-dimensional
image is obtained with high precision with equal or smaller travel
interval (step) of discretely changing the converging beam in an
optical axis direction when an instruction is issued to fetch an
image for a measurement mode in which the sample is measured with
high precision when the brightness information and height
information about the sample are continuously obtained by
discretely performing an iterative operation on the relative
position between a beam condensing position of the object lens and
the sample within the Z scanning area in an optical axis direction
of the converging beam, as compared with a time when the brightness
information and height information about the sample is continuously
obtained by the iterative operation.
25. The method according to claim 23, wherein the Z scanning range
can be changed during the iterative operation on the relative
position between a beam condensing position of the object lens and
the sample within the Z scanning area in an optical axis direction
of the converging beam.
26. The method according to claim 23, wherein a measurement
position is specified on a sample shape continuously updated and
displayed by the iterative operation, and the sample shape is
measured based on the specified measurement position.
27. The method according to claim 23, wherein a 3D image of the
sample shape generated according to the obtained height information
and a confocal image generated according to the brightness
information are continuously updated and displayed with the 3D
image on a same screen by the iterative operation on a same
screen.
28. The method according to claim 27, wherein the confocal image is
an extend image.
Description
CROSS REFERENCE TO RELATED APPLICATIONS
This application claims benefit of Japanese Applications No.
2003-391642, filed Nov. 21, 2003; and No. 2004-328368, files Nov.
12, 2004, the contents of which are incorporated by this
reference.
BACKGROUND OF THE INVENTION
1. Field of the Invention
The present invention relates to a sample information measuring
method and a scanning confocal microscope, and more specifically to
a sample information measuring method and a scanning confocal
microscope for measuring surface information about a height
direction of a sample using the scanning confocal microscope, and
displaying a three-dimensional shape of the sample in a visually
recognizable manner.
2. Description of the Related Art
Conventionally, a scanning confocal microscope applies dotted
illumination to a specimen, converges transmitted light, reflected
light, or fluorescence from the specimen on a confocal diaphragm,
and detects by a photodetector the intensity of the light passing
through the confocal diaphragm, thereby obtaining the surface
information about the specimen. A scanning confocal microscope
scans the surface of the specimen using dotted illumination in
various methods, thereby obtain the surface information about the
specimen in a wide range.
FIG. 1A shows the outline of the configuration of a conventional
scanning confocal microscope.
With the scanning confocal microscope shown in FIG. 1A, a beam
output from a light source 1 passes through a beam splitter 2, and
enters a two-dimensional scanning mechanism 3. The two-dimensional
scanning mechanism 3 has a first optical scanner 3a and a second
optical scanner 3b, performs two-dimensional scanning using
luminous flux, and leads it to an object lens 7. The luminous flux
input to the object lens 7 becomes converging beam and scans the
surface of a sample 8.
The light reflected by the surface of the sample 8 is introduced
from the object lens 7 again to the beam splitter 2 through the
two-dimensional scanning mechanism 3, then reflected by the beam
splitter 2, and converges on a pinhole 10 by an image forming lens
9. The pinhole 10 cuts off the reflected light from the points
other than the beam condensing point of the sample 8 and a
photodetector 11 detects the light only passing through the pinhole
10.
The specimen 8 is held on a sample table 13. A stage 14 and the
photodetector 11 are controlled by a computer 12.
The beam condensing position by the object lens 7 is in a position
optically conjugate with the pinhole 10. When the sample 8 is in
the beam condensing position of the object lens 7, the reflected
light from the sample 8 converges on the pinhole 10 and passes
through the pinhole 10. When the sample 8 is displaced from the
beam condensing position of the object lens 7, the reflected light
from the sample 8 does not converges on the pinhole 10, and does
not pass through the pinhole 10.
FIG. 1B shows the relationship between the relative position (Z) of
the object lens 7 to the specimen 8 and the output (I) of the
photodetector 11.
This relation is called I-Z curve as follows.
As shown in FIG. 1B, when the sample 8 is in the beam condensing
position Z.sub.0 of the object lens 7, the output of the
photodetector 11 indicates a maximum value. As the relative
position of the object lens 7 to the sample 8 leaves from the
position, the output of the photodetector 11 indicates a sudden
decrease.
With the characteristic, if the two-dimensional scanning mechanism
3 performs two-dimensional scanning on the beam condensing point,
and an image is generated by the output of the photodetector 11 in
synchronization with the two-dimensional scanning mechanism 3, then
an image of only a specific height portion of the sample 8 is
formed, and an image (confocal image) is obtained by optically
slicing the sample 8. Furthermore, the sample 8 is discretely moved
on the stage 14 in the optical axis direction, the two-dimensional
scanning mechanism 3 performs scanning in each position to obtain a
confocal image, and the position Z of the stage 14 where the output
of the photodetector 11 indicates the maximum value is detected,
thereby obtaining the height information about the specimen 8.
Additionally, by overlaying and displaying the maximum value of the
output of the photodetector 11 at each point of the sample, an
image can be obtained with all points of the image displayed in
focus (extend image).
When the height of the sample 8 is measured with the
above-mentioned configuration, it is necessary to reduce the amount
of each travel of the stage 14 to enhance the measurement
precision. As a result, it takes some time to make a necessary
measurement. Therefore, a height measuring method is proposed to
enhance the precision in measuring the height of the sample 8
without reducing each the amount of each travel of the stage 14
(refer to Japanese Patent Laid-open Publication No. Hei
9-68413).
In this method, the output of the photodetector 11 is sequentially
obtained while moving the stage 14 based on a predetermined amount
of travel. Then, based on the output of the photodetector 11
relating to the three points, that is, the point indicating the
maximum value of the output and the points before and after the
point indicating the maximum value, an I-Z curve is approximated by
a quadratic curve, and the position of the stage 14 where the
output of the photodetector 11 is to be the maximum is obtained
with the precision equal to or lower than the amount of travel of
the stage 14, thereby obtaining the height information.
There is a disclosed technology of obtaining the surface height
data H(x,y) as the surface information about the sample
corresponding to each pixel based on a confocal image captured at
each height in the height direction of the sample with a view to
measuring the shape of the surface of the sample with high
resolution without reducing the relative amount of travel of the
sample in the height direction (refer to Japanese Patent Laid-open
Publication No. Hei 9-113235).
Practically, the first height position D(m) where the quantity of
light rises to the maximum value in the height direction is
obtained, and the first light quantity Fm(x,y) in the first height
position D(m) and the second light quantity Fm-1(x,y) and the third
light quantity Fm+1(x,y) respectively at the second height position
D(m-1) and the third height position D(m+1) respectively close to
the upper and lower sides of the first height position D(m) are
obtained. Based on these values, a quadratic curve indicating a
change of the quantity of light relative to the height position is
obtained, and the extreme value of the quality of light is obtained
from the quadratic curve. Furthermore, the height position Dmax
corresponding to the extreme value is defined as surface height
data H(x,y).
Additionally, a scanning confocal microscope capable of obtaining
the optical axis direction position and the three-dimensional shape
of a sample without scanning in the optical axis direction is
disclosed. This scanning confocal microscope includes a laser beam
source, a confocal scanner for outputting after passing output
light of the laser beam source through an aperture, an optical
microscope for converging the output light from the confocal
scanner on the sample, a shooting device for shooting the light
passing through the aperture of the confocal scanner in the return
light from the sample, and obtaining a sectional image, and a
control device for obtaining the optical axis direction position of
the sample from the quality of light of the sectional image based
on the optical axis direction position to light quantity
characteristic (Japanese Patent Laid-open Publication No. Hei
11-264933).
SUMMARY OF THE INVENTION
According to an aspect of the present invention, the sample
information measuring method according to the present invention is
used for a scanning confocal microscope, and includes: irradiating
a sample with light from a light source through an object lens;
discretely changing a relative position between the beam condensing
position of the object lens and the sample along an optical axis
direction of the converging beam; obtaining light intensity
information from the sample in each relative position; extracting
plural pieces of light intensity information from a group of the
plural pieces of light intensity information; estimating a maximum
value in a change curve matching the plural pieces of extracted
light intensity information and the corresponding relative
positions; and obtaining a maximum value of the estimated light
intensity information and the relative position respectively as
brightness information and height information. The method
continuously obtains the brightness information and the height
information about the sample by discretely and iteratively
performing a reciprocal operation in the optical axis direction of
the converging beam on the relative position between the beam
condensing position of the object lens and the sample.
BRIEF DESCRIPTION OF THE DRAWINGS
The present invention will be more apparent from the following
detailed description when the accompanying drawings are
referenced.
FIG. 1A shows the outline of the configuration of the conventional
scanning confocal microscope;
FIG. 1B shows the relationship between the relative position (Z) of
the object lens 7 and the sample 8 and the output (I) of the
photodetector 11;
FIG. 2A shows the configuration of the scanning confocal microscope
to which the first embodiment of the present invention is
applied;
FIG. 2B is an explanatory view of an example of the shape of a
sample to be measured;
FIG. 3 shows an example of an image display when a focal point is
obtained on the plane c;
FIG. 4 is an explanatory view of setting a scanning range in the X
direction;
FIG. 5 shows an I-Z curve of an optical intensity group of the
points on the planes a, b, and c;
FIG. 6 shows an optical intensity group in the positions of Z(-2)
through Z(2);
FIG. 7 shows an example of an image display when a focal point is
obtained on the entire plane;
FIG. 8 shows an example of a display according to the second
embodiment of the present invention;
FIG. 9 shows an example of a display of the distance between two
indicated points;
FIG. 10 shows an image of an example indicating two arbitrary
points;
FIG. 11 shows examples of displays of .DELTA.X, .DELTA.Y, and
.DELTA.Z between the two indicated points;
FIG. 12 shows an example (1) of displaying a 3D image 21 and an
extend image 22 on a monitor 15;
FIG. 13 shows an example (2) of displaying a 3D image 21 and an
extend image 22 on a monitor 15;
FIG. 14 shows an example (3) of displaying a 3D image 21 and an
extend image 22 on a monitor 15;
FIG. 15 shows an example (4) of displaying a 3D image 21 and an
extend image 22 on a monitor 15;
FIG. 16 shows an example (5) of displaying a 3D image 21 and an
extend image 22 on a monitor 15;
FIG. 17 shows an example (6) of displaying a 3D image 21 and an
extend image 22 on a monitor 15;
FIG. 18 shows the configuration of the scanning confocal microscope
to which the fifth embodiment of the present invention is
applied;
FIG. 19 shows an example (1) of a 3D image 31 and a non-confocal
image 32 displayed on the monitor 15;
FIG. 20 shows an example (2) of a 3D image 31 and a non-confocal
image 32 displayed on the monitor 15;
FIG. 21 shows an example (3) of a 3D image 31 and a non-confocal
image 32 displayed on the monitor 15;
FIG. 22 shows an example (4) of a 3D image 31 and a non-confocal
image 32 displayed on the monitor 15;
FIG. 23 shows an example (1) of a 3D image 31 and a non-confocal
image 33 displayed on the monitor 15;
FIG. 24 shows an example (2) of a 3D image 31 and a non-confocal
image 33 displayed on the monitor 15;
FIG. 25 shows an example (3) of a 3D image 31 and a non-confocal
image 33 displayed on the monitor 15;
FIG. 26 shows the configuration of the scanning confocal microscope
to which the sixth embodiment of the present invention is
applied;
FIG. 27 shows an example of displaying a 3D image 41 and a color
image 42 on the monitor 15;
FIG. 28 is a flowchart shoring the flow of the sample information
measuring process; and
FIG. 29 shows an example of displaying a three-dimensional image
update display lamp showing an iterative display timing of the 3D
image 31.
DESCRIPTION OF THE PREFERRED EMBODIMENTS
The embodiments of the present invention are described below by
referring to the attached drawings.
FIG. 2A shows the configuration of the scanning confocal microscope
to which the first embodiment of the present invention is
applied.
In the scanning confocal microscope shown in FIG. 2A, the light
output from the light source 1 passes through the beam splitter 2,
and is then input to the two-dimensional scanning mechanism 3. The
two-dimensional scanning mechanism 3 comprises the first optical
scanner 3a and the second optical scanner 3b, performs scanning
using luminous flux in a two-dimensional manner, and leads the
luminous flux to the object lens 7. The luminous flux input to the
object lens 7 scans the surface of the sample 8 as converging
beams.
The light reflected by the surface of the sample 8 is led to the
beam splitter 2 through the two-dimensional scanning mechanism 3
from the object lens 7, and then reflected by the beam splitter 2,
and converges at the pinhole 10 by the image forming lens 9. The
pinhole 10 cuts off the reflected light from the portions other
than the beam condensing point of the sample 8, and only the light
passing through the pinhole 10 is detected by the photodetector
11.
A Z revolver 6 has a plurality of object lens 7, inserts the object
lens 7 of desired power into an optical path of two-dimensional
scanning, moves in the Z axis direction, and changes the relative
position between the beam condensing position of the object lens 7
and the sample 8.
The sample 8 is placed on the sample table 13, and can be moved in
the XY directions by the stage 14. The two-dimensional scanning
mechanism 3, Z revolver 6, and photodetector 11, etc. are
controlled by a microscope control program stored in the computer
12, and the user can operate each unit on the operation screen
displayed on a monitor 15.
The beam condensing position by the object lens 7 is conjugate with
the pinhole 10. When the sample 8 is in the beam condensing
position by the object lens 7, the reflected light from the sample
8 converges on the pinhole 10, and passes through the pinhole 10.
When the sample 8 is displaced from the beam condensing position,
the reflected light from the sample 8 does not converge on the
pinhole 10, and does not pass through the pinhole 10.
According to the I-Z curve indicating the relationship between the
relative position (Z) of the object lens 7 shown in FIG. 1B to the
sample 8 and the output (I) of the photodetector 11, when the
sample 8 is in the beam condensing position Z.sub.0 of the object
lens 7, the output of the photodetector 11 indicates the maximum
value, and as the relative position between the object lens 7 and
the sample 8 deviates from the position, the output of the
photodetector 11 suddenly drops.
By the above-mentioned characteristic, only the specific height of
the specimen 8 is displayed as an image, and an image can be
obtained as an optically sliced image (confocal image) of the
sample 8 when the two-dimensional scanning is performed on the beam
condensing point and the output of the photodetector 11 is
displayed as an image in synchronization with the two-dimensional
scanning mechanism 3. Then, the image is displayed with the
operation screen on the monitor 15.
The sample information measuring method to which the present
invention is applied is explained using the scanning confocal
microscope shown in FIG. 2A.
FIG. 2B is an explanatory view showing an example of the shape of a
sample to be measured.
The sample shown in FIG. 2B is assumed as the sample 8 to be
measured by the scanning confocal microscope. That is, assume that
the sample 8 has three surfaces having different heights (thickness
in the Z direction), that is, the surfaces a, b, and c from one end
to the other end.
FIG. 3 shows an example of displaying an image when a focus is
obtained on the surface c.
First, the two-dimensional scanning of the scanning confocal
microscope is started, and the focus is obtained on the surface c
of the sample 8. At this time, the monitor 15 displays the image as
shown in FIG. 3. Practically, the scanning confocal microscope
starts obtaining an image using a "scanning start/stop" button, the
object lens 7 of desired power is selected by a "object" button,
and an adjustment is made such that an observed portion can be
displayed in a desired size together with the "zoom" scroll bar.
Then, the Z revolver 6 is moved up and down using the "position Z"
scroll bar, and the focal surface of the object lens 7 is aligned
with the surface c of the sample 8.
FIG. 4 is an explanatory view of setting the scanning range in the
Z direction.
After the focal surface is aligned with the surface c of the sample
8, the scanning range in the Z direction is determined. The
condition of the scanning range in the Z direction is set with the
target of the area indicated by the "Z scanning range" shown in
FIG. 4. The conventional scanning confocal microscope estimates and
determines the rough shape of the sample 8 and the area of the "Z
scanning range" while checking the two-dimensional image by moving
the focal position up and down. However, the present invention can
set the Z scanning range from the display of the sample 8 directly
displayed on the monitor 15 by the operation of the process
described below. On the screen, the reference view range in the XY
direction is selected using the "object" button, and the reference
position in the Z direction is selected using the "position Z"
scroll bar. Then, the three-dimensional scanning range is set using
the "zoom" and "Z range" scroll bars.
The computer 12 stores the brightness and height arithmetic
program. By executing the brightness and height arithmetic program,
the brightness and height information using a confocal image can be
obtained.
Described below is the brightness and height measuring process.
When the Z range scroll bar is set to a value other than zero (0),
the Z revolver 6 starts moving stepwise up and down in the Z
scanning range corresponding to the set value using the scroll bar
with the current focal position (surface c) set in the center as
shown in FIG. 4. While checking the image of the sample 8 displayed
on the monitor 15, the user sets a desired Z scanning range on the
operation screen. The Z revolver 6 moves at a predetermined travel
pitch .DELTA.Z in the set Z scanning range, and a confocal image is
obtained for each Z relative position. For simple explanation, it
is assumed that the number of obtained confocal images is 5, that
is, the frequency of the travel of the Z revolver 6 is 4 to the
positions Z(-2), Z(-1), Z(0), Z(1), and Z(2). The light intensity
information about the arbitrary point (for example, the points on
the surfaces a, b, and c) of the sample 8 at this time is
obtained.
FIG. 5 shows an I-Z curve of a light intensity point group on the
surfaces a, b, and c. FIG. 6 shows the light intensity point group
in the positions of the Z(-2) through Z(2).
The light intensity points group on the surfaces a, b, and c
indicate the values on the I-Z curve as each point shown in FIG.
5.
Then, the light intensity information on each point is compared
with each other, and (I(n), Z(n)) indicating the maximum intensity,
and the values before and after the maximum intensity point
(I(n-1), Z(n-1)), (I(n+1), Z(n+1)) are extracted. Relating to the
surface a in the case shown in FIG. 4, the maximum intensity point
is (Ia(-1), Za(-1)), and the values before and after the point are
(Ia(0), Za(0)), (Ia(-2), Za(-2)). By assuming the approximate
quadratic curve passing through the three points, and obtaining the
extreme values, the true maximum value Ia[max] and the position
Za[max] of the Z revolver 6 for the maximum value can be obtained.
Therefore, the brightness and relative height of the surface of the
sample 8 can be obtained with the resolution of the travel pitch
.DELTA.Z or higher. The same holds true with the surfaces b and c.
The maximum intensity point on the surface b is (Ib(1), Zb(1)), and
the points before and after the maximum intensity point are (Ib(0),
Zb(0)), (Ib(2), Zb(2)). Therefore, the values Ib [max] and Zb[max]
are obtained. The maximum intensity point for the surface c is
(Ic(0), Zc(0)), and the points before and after are (Ic(-1),
Zc(-1)), (Ic(1), Zc(1)), thereby obtaining the values Ic[max] and
Zc[max].
FIG. 7 shows an example of displaying an image when a focus is
obtained on the entire image.
Since brightness and height information can be extracted by five
confocal images, the stepwise travel of the Z revolver 6 is
repeated in the order of
Z(-2).fwdarw.Z(-1).fwdarw.Z(0).fwdarw.Z(1).fwdarw.Z(2).fwdarw.Z(1).fwd-
arw.Z(0).fwdarw.Z(-1).fwdarw.Z(-2).fwdarw. . . . so that the
structure of the sample 8 from the surface a to the surface b can
be obtained. At this time, if the position in the height direction
or the width displayed in the microscopic image window is not
appropriate, then the settings can be optimally adjusted while
checking the extend image by changing the center of the Z scanning
range, that is, the "position Z", or the "range Z" itself
respectively. This means if the image for which the focus is
entirely obtained is displayed in the microscopic image window,
then it is determined that the scanning is being operated in a
sufficient area in the height direction of the sample 8.
On the other hand, about 1.8 second is required to obtain five
images because about 200 ms is required to obtain one confocal
image and about 200 ms is required for each stepwise travel of the
Z revolver 6 by assuming that the image size is 1024.times.768, and
the Z scanning range is 5 .mu.m (.DELTA.Z=1 .mu.m). With the
additional two seconds as processing time, a total of two seconds
are enough to update the brightness and height information.
However, since images are continuously obtained by moving the Z
revolver 6 up and down, the fifth image can be used as the first
confocal image in the next process of obtaining the brightness and
height information. Therefore, one updating operation can be
completed within two seconds. This is a sufficiently practical
updating speed. Since various combinations are made by the number
of lines of one confocal image, the amount of stepwise travel of
the Z revolver 6, and the number of steps (range) for an update
speed, a user can optionally select a desired combination from
among predetermined combinations. For example, when an updating
operation is to be quickly performed, the number of lines of a
confocal image is to be limited, and the Z scanning range is to be
narrowed to a minimal level. In this embodiment, the number of
images is five, but three images can be used in obtaining the
maximum value in principle from the plural pieces of light
intensity information, thereby further raising the updating
speed.
Described below is the second embodiment of the present
invention.
The configuration of the scanning confocal microscope according to
the second embodiment is the same as the configuration of the
embodiment explained above by referring to FIG. 2A. However, the
information about the sample 8 displayed on the monitor 15 is
represented in a three-dimensional array.
FIG. 8 shows an example of a display according to the second
embodiment of the present invention.
The display example shown in FIG. 8 refers to simultaneously
displaying combined brightness information about the surfaces of
the sample 8 on the surface portion of the height information
obtained by the brightness and height arithmetic program. Since it
is constantly updated in synchronization with the laser scanning of
the scanning confocal microscope, it is a true representation in a
three-dimensional array of the actual sample 8 in comparison with
the two-dimensional display by an extend image. With this display,
the user can obtain at a glance the detailed three-dimensional
information about the sample 8. The function of rotation,
enlargement, reduction, etc. provided for general 3D display
software can be arbitrarily performed, and the user can observe an
object at a desired angle.
Described below is the third embodiment of the present
invention.
As described above, since three-dimensional information about the
sample 8 is continuously obtained and updated, it is also possible
to continuously measuring any optional unit.
FIG. 9 shows an example of displaying the distance between two
indicated points.
It is also possible to carry out a measurement directly using a 3D
display. Measurement results can be continuously obtained with the
update of an image by specifying the measurement position using a
line cursor from the sample 8 displayed in the three-dimensional
array, and specifying two points in a profile window.
There is, for example, a mouse, etc. connected to the computer
12.
For example, as shown in FIG. 10, the "step" in the measurement
items is selected, and the line cursor is moved on the target point
of the extend image (cross cursor 1 and cross cursor 2). Since the
sectional profile is obtained on the line, two points are specified
in the profile. Then, the steps between the points are continuously
measured, and the display of the measured value is updated in
synchronization with the drawing. While the conventional scanning
confocal microscope fetches a three-dimensional image for a
measurement, the flow of the operations terminates and the process
is performed again from the beginning on another condition, the
function according to the present invention can continuously
perform the processes of "setting a condition".fwdarw."checking by
measurement".fwdarw."amending the condition".
FIG. 11 shows an example of displaying .DELTA.X, .DELTA.Y, and
.DELTA.Z between two points.
Since it is hard to specify one point in the space on the sample 8
displayed in the three-dimensional array using a mouse, an
arbitrary one point can be specified by displaying a cross cursor
displayed in the profile on a three-dimensional image. This can be
a profile measurement in one line, or the specification can be made
among a plurality of different profiles. A measurement result
refers to .DELTA.X, .DELTA.Y, and .DELTA.Z between two specified
points, the distance between two points, etc. continuously measured
in synchronization with the scanning, and the display is also
updated.
Described below is the fourth embodiment of the present
invention.
The scanning confocal microscope to which the fourth embodiment of
the present invention is applied is the same as the scanning
confocal microscope to which the above-mentioned first through
third embodiments are applied.
That is, the scanning confocal microscope to which the fourth
embodiment of the present invention is applied generates an extend
image indicating a focus obtained on the entire image and a height
map image. The height map image can be processed by the computer 12
and displayed on the monitor 15 in a three-dimensional array.
The scanning confocal microscope to which the fourth embodiment of
the present invention is applied displays an extend image together
with the 3D image on the same screen while continuously performing
update.
FIGS. 12 through 17 show an example of displaying the 3D image 21
and the extend image 22 displayed on the monitor 15. The 3D image
21 indicating the three-dimensional shape of the surface of the
sample 8 and the extend image 22 having the focus obtained on the
entire image are displayed on the monitor 15 and simultaneously and
continuously updated.
FIG. 12 shows an example of displaying the three-dimensional image
21 and the two-dimensional extend image 22 in the same screen size.
FIG. 13 shows an example of reducing the two-dimensional extend
image 22 on a part of the three-dimensional image 21 displayed on
the full screen. It is more preferable if ratio of the display size
can be arbitrarily changed so that the user can easily see the
display.
FIGS. 14 through 16 show an example of reducing the two-dimensional
extend image 22 on a part of the three-dimensional image 21
displayed on the full screen. FIG. 14 shows an example of
representing the height information in gray scales as the
three-dimensional image 21 displayed on the full screen. FIG. 15
shows an example of displaying the height information as the
three-dimensional image 21 displayed on the full screen using
lines. FIG. 16 shows an example of displaying the height
information as a three-dimensional image 21 displayed on the full
screen using a mesh.
A displayed two-dimensional image can be the extend image 22
treated in predetermined image processing. For example, if an edge
extraction filter is applied to the extend image 22 and the
resultant image is displayed, a three-dimensional shape can be
observed using the 3D image 21, and the edge portion can be
simultaneously observed using the extend image 22. As image
processing, for example, general image processing methods such as
binarizing filtering, boundary line extraction filtering, etc. can
be applied. The image processing can also be performed during
display in a continuously updating process.
Additionally, the extend image 22 displayed as a two-dimensional
image can be represented as a height map image as shown in FIG. 17,
or a contour line image based on the height map image. Thus, the
three-dimensional image 21 and the extend image 22 indicating the
two-dimensional height information are simultaneously displayed,
and continuously updated with a lapse of time, thereby allowing the
user to easily recognize the information in the height
direction.
The 3D image 21 can arbitrarily rotate, enlarge, or reduce an image
during operation, and the Z scanning range set as the current lower
limit of the focal position can be moved up and down. Thus, the
first Z scanning range can be easily specified for the sample 8
whose amount of step is predetermined based on the designed value,
etc.
Thus, by continuously updating and displaying the 3D image 21 and
the extend image 22 of the sample 8 on the same screen of the
monitor 15, the user can simultaneously observe the
three-dimensional information and the two-dimensional information
about the sample 8. Therefore, the user can visually observe the
surface status of the sample 8 easily.
Described below is the fifth embodiment of the present
invention.
FIG. 18 shows the configuration of the scanning confocal microscope
to which the fifth embodiment of the present invention is
applied.
The scanning confocal microscope to which the fifth embodiment of
the present invention is applied compares further comprises a half
mirror 16 and an photodetector 17 as compared with the scanning
confocal microscope to which the first embodiment of the present
invention is applied.
That is, the light reflected by the surface of the sample 8 is
introduced from the object lens 7 again to the beam splitter 2
through the two-dimensional scanning mechanism 3, converges by the
image forming lens 9, and is divided by the half mirror 16 and
detected by the photodetector 17 and the photodetector 11 through
the pinhole 10. The image detected by the photodetector 17 is a
non-confocal image and has a large depth of focus.
The scanning confocal microscope to which the fifth embodiment of
the present invention is applied simultaneously displays a
non-confocal image together with the three-dimensional image.
Then, the sample information measuring method using the scanning
confocal microscope shown in FIG. 18 is explained below. The method
of obtaining a height map image and repeatedly displaying a
three-dimensional image is the same as those according to the
above-mentioned embodiments.
FIGS. 19 through 22 show an example of displaying the 3D image 31
and the non-confocal image 32 on the monitor 15. The
three-dimensional image 31 and the non-confocal image 32 of the
surface of the sample 8 are simultaneously and continuously
displayed on the monitor 15.
At this time, the display of the 3D image 31 is updated each time
the height information is extracted, and the display of the
non-confocal image 32 is updated for each position Z. That is, when
the operation as shown in FIG. 4 is performed, the display of the
3D image 31 is updated each time a travel from Z (-2) to Z (2) is
detected, and the display of the non-confocal image 32 is updated
at each position of Z (-2), Z (-1), Z (0), Z(1), and Z(2).
FIG. 19 shows an example of displaying the 3D image 31 and the
non-confocal image 32 at Z (-1) on the monitor 15. FIG. 20 shows an
example of displaying the 3D image 31 and the non-confocal image 32
at Z(0) on the monitor 15. FIG. 21 shows an example of displaying
the 3D image 31 and the non-confocal image 32 at Z(1) on the
monitor 15. FIG. 22 shows an example of displaying the 3D image 31
and the non-confocal image 32 at Z(2) on the monitor 15.
As shown in these FIGS. 19 through 22, the display of the 3D image
31 is updated each time the height information is extracted.
Therefore, the same image is displayed from FIG. 19 to FIG. 21, and
the image is first updated in FIG. 22. However, since the display
of the non-confocal image 32 is updated for each position Z, the
image is updated in each of FIGS. 19 through 22. As in the fourth
embodiment of the present invention, the non-confocal image 32
displayed in this case can be in the reduced image displaying
method.
Thus, by simultaneously and continuously updating and displaying
the non-confocal image 32 obtained at each position Z in the Z
scanning range on the screen of the same monitor 15 as the 3D image
31, losing the observation place of the sample 8 by the user when
the sample 8 is moved in the XY directions can be avoided.
Especially since the non-confocal image 32 has a large depth of
focus, the information about the surface of the sample 8 can be
easily obtained at any position Z, and the user can easily adjust
the position of the sample 8 while simultaneously seeing the
non-confocal image 32 and the 3D image 31. The more preferable
operability can be obtained by the optical scanners 3a and 3b
continuously during the scanning in the range Z, and updating the
non-confocal image 32 for each frame.
Furthermore, the image displayed as a two-dimensional image can be
a confocal image at each position Z instead of the non-confocal
image 32. In this case, the depth of focus becomes smaller, but the
change transition of the portions for which a focus can be obtained
can be observed simultaneously with the three-dimensional
shape.
FIGS. 23 through 25 show an example of the 3D image 31 and the
non-confocal image 33 displayed on the monitor 15. That is, the 3D
image 31 indicating the three-dimensional shape and the
non-confocal image 33 of the surface of the sample 8 are
simultaneously updated and displayed on the monitor 15.
FIG. 23 shows an example of displaying the 3D image 31 and the
non-confocal image 33 on the monitor 15 at Z (-1). FIG. 24 shows an
example of displaying the 3D image 31 and the non-confocal image 33
on the monitor 15 at Z (0). FIG. 25 shows an example of displaying
the 3D image 31 and the non-confocal image 33 on the monitor 15 at
Z (1).
Described below is the sixth embodiment of the present
invention.
FIG. 26 shows the configuration of the scanning confocal microscope
to which the sixth embodiment of the present invention is
applied.
The scanning confocal microscope to which the sixth embodiment of
the present invention further comprises the white light source 19
and a color detector 20 as compared with the scanning confocal
microscope to which the first embodiment shown in FIG. 2A is
applied.
That is, the light reflected by the surface of the sample 8 forms
an image on the color detector 20 such as a color CCD, etc., a
signal captured by the color detector 20 is fetched by a color
image fetch board, and the color image is displayed on the monitor
15 together with the three-dimensional image.
The sample information measuring method using the scanning confocal
microscope shown in FIG. 26 is explained below. The method for
obtaining a height map image and repeatedly displaying a
three-dimensional image is the same as those according to the
above-mentioned embodiments.
FIG. 27 shows an example of displaying the 3D image 41 and the
color image 42 on the monitor 15. The three-dimensional image 41
and the color image 42 of the surface of the sample 8 are
simultaneously and continuously updated and displayed on the
monitor 15.
At this time, the display of the 3D image 41 is updated each time
the height information is extracted, and the Z moving operation,
the scanner scanning, and the capturing and drawing the color image
42 can be asynchronously performed. Therefore, the color image 42
can be updated substantially at the frame rate.
As described above, by continuously updating and displaying the
color image 42 obtained from the color detector 20 together with
the 3D image 41 on the screen of the same monitor 15, losing the
observation place of the sample 8 by the user when the sample 8 is
moved in the XY directions can be avoided. Furthermore, since the
information about the color can be obtained by the color image 42,
the user can easily recognize the status of the surface of the
sample 8, and can easily adjust the position of the sample 8 while
simultaneously watching the color image 42 and the 3D image 41.
Instead of the display and update of the color image 42 at the
frame rate, the color extend image whose focus has been composed
such that the focus can be obtain on the entire image according to
the information about the contrast, etc. from the information about
the color image 42 can be displayed and updated each time the Z
scanning is performed. In this case, a color extend image can be
attached to the surface of the 3D image 41 in place of the extend
image configured by the confocal image.
Explained below is the flow of the sample information measuring
method common among the above-mentioned embodiments.
FIG. 28 is a flowchart of the flow of the sample information
measuring method.
First, in step S281, the user confirms the sample 8 using a
two-dimensional image by the scanning confocal microscope (LSM). In
step S282, a start button is pressed to perform repeated display in
the three-dimensional array.
In step S283, the Z position start button is operated and the
observation position of the sample 8 to be observed in the Z
direction. In step S284, the Z scanning range (width from the upper
end position to the lower end position) of the three-dimensional
repeated display is adjusted.
Then, the scanning confocal microscope according to each embodiment
displays the 3D images 21, 31, and 41 by repeatedly scanning the
sample 8 in the Z direction.
At this time, for example, as shown in FIG. 29, when the 3D image
update indicator lamps 23 and 24 indicating the iterative display
timing of the 3D image 21 are displayed on the monitor 15, the user
can be informed of the timing of the update of the 3D image 21. The
display example shown in FIG. 29 shows an example of the alternate
display of the 3D image update indicator lamps 23 and 24 for each
cycle at which the 3D image 21 is displayed. That is, at the first
cycle, the 3D image update indicator lamp 23 is turned on while the
3D image update indicator lamp 24 is turned off. At the second
cycle, the 3D image update indicator lamp 23 is turned off while
the 3D image update indicator lamp 24 is turned on. At the third
cycle, the 3D image update indicator lamp 23 is turned on again and
the 3D image update indicator lamp 24 is turned off.
As described above, at each cycle of the display of the 3D image
21, the 3D image update indicator lamps 23 and 24 are displayed,
thereby the user can be informed of the update timing of the 3D
image 21 and the change timing of the measurement parameter. The 3D
image update display indicating the timing of the iterative
scanning display is not limited to the above-mentioned application,
but one indicator lamp can be used, or various graphic or bar type
level meters, etc. can be used.
It returns to the explanation of FIG. 28.
In step S285 shown in FIG. 28, the user determines whether or not
the displayed 3D images 21, 31, and 41 have expected
three-dimensional shapes. If the user determines that they do not
have expected three-dimensional shapes (NO in step S286), then the
processes in and after step S283 are repeated, thereby re-adjusting
the observation position of the sample 8 to be observed in the Z
direction (step S283) and also re-adjusting the Z scanning range of
the three-dimensional iterative display (step S284).
When it is determined in step S286 that the image has an expected
three-dimensional shape (YES in step S286), the three-dimensional
image fetch button for fetching the image information in the
measurement mode is pressed in step S287, thereby stopping the
three-dimensional iterative display. In step S288, the
high-precision three-dimensional image automatically converted to
an equal or smaller size as compared with the Z scanning step or
the XY scanning step used when observation is made with the
three-dimensional iterative display in the same scanning range as
in the observation with the three-dimensional iterative display in
step S285 is obtained. Therefore, the measurement higher in
precision than the measurement of the image continuously updated
and displayed can be performed.
Thus, each embodiment according to the present invention is
described above, but the configuration of the scanning confocal
microscope according to the present invention is not limited to the
configuration shown in FIG. 2A, 18, or 26, and can be applied to
various types of scanning confocal microscopes.
For example, a configuration of rotating a Nipkow disk at a high
speed with a plurality of fine apertures designed in a spiral
pattern on a disk can be used. At this time, the Nipkow disk also
functions as the fine apertures arranged in the positions conjugate
with the beam condensing position of the object lens, and a
two-dimensional image sensor such as a CCD, etc. is used as an
optical detector. Furthermore, a two-dimensional optical scanning
mechanism is replaced with a one-dimensional optical scanner in
scanning one line of a sample using a converging beam of an object
lens and measuring the sectional shape of the sample.
As a moving mechanism for relatively moving the beam condensing
position of the object lens 7 with the position of the sample 8,
the Z revolver 6 for moving the object lens 7 can be replaced with
a stage mechanism for moving the position of the sample 8.
An applicable configuration is not limited to those described
above, but various types of scanning confocal microscopes can be
used. That is, the scanning confocal microscope and the sample
information measuring method according to the present invention can
be designed in various configurations or shapes without limitations
in the range of the gist of the present invention.
* * * * *